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United States Patent |
6,043,213
|
Tsubota
|
March 28, 2000
|
Drug composition comprising albumin as active ingredient
Abstract
The present invention provides a pharmaceutical composition for treatment
of corneal and conjunctival lesion, and dry eye comprising albumin as an
active ingredient. The pharmaceutical composition is also useful for
increase of eye surface epithelium mucin secretion. The present invention
further provides a method for treatment of corneal and conjunctival
lesion, and dry eye, which comprises administering, to a subject in need
of such treatment, albumin in an amount effective. In addition, the
present invention provides a use of albumin for manufacture of a
pharmaceutical composition of the present invention.
Inventors:
|
Tsubota; Kazuo (Funabashi, JP)
|
Assignee:
|
R-Tech Ueno, Ltd. (Osaka, JP)
|
Appl. No.:
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981229 |
Filed:
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December 19, 1997 |
PCT Filed:
|
April 17, 1997
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PCT NO:
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PCT/JP97/01329
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371 Date:
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December 19, 1997
|
102(e) Date:
|
December 19, 1997
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PCT PUB.NO.:
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WO97/39769 |
PCT PUB. Date:
|
October 30, 1997 |
Foreign Application Priority Data
| Apr 19, 1996[JP] | 8-098090 |
| Apr 26, 1996[JP] | 8-106866 |
Current U.S. Class: |
514/2; 514/912 |
Intern'l Class: |
A61K 037/00 |
Field of Search: |
514/2,912
|
References Cited
U.S. Patent Documents
4775531 | Oct., 1988 | Gilbard.
| |
Foreign Patent Documents |
5-310592 | Nov., 1993 | JP.
| |
6-271478 | Sep., 1994 | JP.
| |
93-64188 | Sep., 1994 | JP.
| |
Other References
Chemical Abstracts 122: 38840. Tsuboto et al, 1994.
Database Drugu, AN 84-16297. Koning et al. Combination Therapy for
Dendritic Keratitis with Acyclovir and Alpha--Interferon. Arch.
Ophthalmol. 101(12), 1866-68 (Dec. 1983).
Gupta et al. Trypsin and Serum Albumin in Tear Fluids in Acute Adenovirus
Conjunctivitis. BR. J. Ophthalmol 72(5), 390-393 (May 1988).
Pedersen et al. The origin of immunoglobulin G in bovine tears. Acata Path.
Microbiol.Scand.Set.B, 81B/2, 245-252 (Feb. 1973).
Zavaro et al. Proteins in Tears from Healthy and Diseased Eyes. Symposium
on Uveal Melanomes, Nov. 24, 1979. Doc Ophthalmol 50(1) (Recd. 1981).
185-199 (Jan. 1981).
Ran S et al. Topical Plasma Fibronectin Compared to Albumin and Saline for
Wond Healing of Denervated Corneal Epithelial Defeacts. Annual Spring
Meeting of Association for Research in Vision and Ophthalmology,
Sarasotata, Florida, USA, Apr. 3. (Mar. 1989).
|
Primary Examiner: Fay; Zohreh
Attorney, Agent or Firm: Sughrue, Mion, Zinn Macpeak & Seas, PLLC
Parent Case Text
This application is a continuation of U.S. Ser. No. 08/752,928, filed Nov.
21, 1996 and U.S. Ser. No. 08/752,941, filed Nov. 21, 1996, and embodies
the subject matter disclosed therein.
Claims
What is claimed is:
1. A method for treatment of dry eye which comprises administering, to a
subject in need of such treatment, albumin in an amount effective for
treatment of dry eye.
2. The method according to claim 1, wherein the said albumin is human
origin albumin.
3. The method according to claim 1, wherein the said albumin is
administered in the form of eye drops.
4. The method according to claim 1, which comprises administering a
pharmaceutical composition containing 1 to 1000 mg/ml of albumin and
pharmaceutically acceptable carrier.
5. A method for increasing eye surface epithelium mucin secretion, which
comprises administering, to a subject in need of such administration,
albumin in an amount effective for increasing eye surface epithelium mucin
secretion.
6. The method according to claim 5, wherein the said albumin is human
origin albumin.
7. The method according to claim 5, wherein the said albumin is
administered in the form of eye drops.
8. The method according to claim 5, which comprises administering a
pharmaceutical composition containing 1 to 1000 mg/ml of albumin and
pharmaceutically acceptable carrier.
Description
TECHNICAL FIELD
The present invention provides a pharmaceutical composition comprising
albumin as an active ingredient. The composition of the present invention
is useful for treatment of the conditions such as corneal and conjunctival
lesion and dry eye. The composition of the present invention has an
ability to increase eye surface epithelium mucin secretion and also has an
ability to diffuse oil. The present invention further provides methods for
treatment of corneal and conjunctival lesion, treatment of dry eye
conditions, and increase of eye surface epithelium mucin secretion using
the composition.
BACKGROUND ART
Corneal and conjunctival lesion is caused by forming defects from the
surface to epithelium. The cause may include pathogenic factors such as
keratoconjunctivitis sicca (dry eye), various keratoconjunctivitis,
allergy and infection of microorganisms (e.g. virus, bacteria, fungus,
etc.), chemical factors such as cytotoxicity by chemicals and corrosion
due to acid and alkaline, physical factors such as xerophthalmia, injury
due to foreign matter (e.g. contact lens, etc.) and hot water, and the
like. It has recently been reported that antiseptics contained in an
ophthalmic composition (e.g. benzalkonium chloride, chlorobutanol, etc.)
and ophthalmic agents (e.g. aminoglycoside antibiotics, non-steroidal
anti-inflammatory drugs, IDU, pimaricin, etc.) cause a lesion of corneal
epithelium (ectocornea).
For the present, in order to treat the corneal and conjunctival lesion,
chondroitin sulfate, glutathione, hyaluronic acid, fibronectin, EGF, and
the like are administered or an artificial tear solution is also
administered for the purpose of replenishing a tear solution, but the
effect of these treatments is not yet sufficient.
Dry eye is defined as a condition with decrease or change in quality of
tear, irrespective of the presence or absence of corneal and conjunctival
lesion (Yamada et al. GANKI 43, 1289-1293(1992)). There are various
factors for causing the dry eye, but no suitable method to recover the
decreased amount of tear normal has been found yet.
For the present, in order to treat the dry eye, an artificial tear solution
for the purpose of replenishing tear, and chondroitin sulfate,
glutathione, hyaluronic acid, fibronectin, EGF, and the like for the
purpose of relieving subjective symptoms are administered, but the effects
are not yet sufficient.
From the recent investigation, it is believed that normal eye surface
epithelium expresses mucin-like glycoproein and that said glycoprotein
takes an active part in maintaining tear film. Furthermore, mucin secreted
from conjunctival germ cells has been known to be responsible to stabilize
tear film. So, defect of eye surface epithelium caused by any factor may
induce abnormal eye mucin secretion and thereby unstable tear film. The
unstable tear film may lower the interaction level between the epithelium
and the tear film, and thereby the lesion of corneal and conjunctival
epithelium may become worse (Norihiko Yokoi in "GANKA CHIRYO NO KOTSU TO
OTOSHIANA" (Techniques for treatment in Ophthalmologic field), edited by
FUMIO KOGURE, published by KABUSHIKI KAISHA NAKAYAMA SHOTEN, Tokyo, Japan
pages 26-27 (1995)). From these points of view, it is suggested that
conditions such as corneal and conjunctival lesion or dry eye can be
treated if the tear film is stabilized by increasing mucin secretion or
any other mean.
Albumin is a protein that exists widely in an animal/vegetable tissue or a
body fluid, such as human serum, tear solution. For example, the human
serum albumin is used for treating hypoalbuminemia, hemorrhagic shock, and
the like. In the ophthalmic field, it is also known to use as a stabilizer
for protein preparations such as fibronectin or interferon. It has been
proposed to use the preparations such as fibronectin and interferon for
treating corneal lesion (Japanese Patent Kokai No. Sho 61-103838 and No.
Hei 6-271478), but there is no knowledge that albumin itself is effective
for treating corneal and conjunctival lesion or dry eye.
DISCLOSURE OF INVENTION
The present invention provides a pharmaceutical composition for
ophthalmology comprising albumin as an active ingredient. The present
composition is useful for treating corneal and conjunctival lesion. The
present composition also useful for treating dry eye.
The origin of albumin used for the composition of the present invention is
not specifically limited. When albumin has an antigenicity, however, a
problem such as allergy arises and, therefore, it is not preferred. Human
origin albumin, e.g. human serum albumin is preferably used.
Human serum albumin, which is purified to the purity suitable for using
normally in medical applications, can preferably be used in the present
invention without any particular problem. That is, those containing not
less than 80% of albumin (in case of analyzing with the electrophoresis)
are preferred. In order to inactivate virus, etc., those obtained by
heat-treating are preferred. Particularly, human serum albumin, which is
commercially available as a drug, is preferably used.
Albumin produced by microorganism obtained by gene recombination is also
preferably used in the present invention. The manufacturing method
according to the gene recombination technique is well known to persons
skilled in the art. Briefly explaining, a vector containing a gene coding
a desired albumin (e.g. human albumin) may be introduced into a host cell
to transform it. The transformed cell producing the desired protein may be
selected and cultured, then, human albumin may be isolated and purified
from the cultured supernatant or cell extract. Examples of the host cell
include yeast, Escherichia coli, and the like, which are used ordinary by
persons skilled in the art so as to produce a protein. In view of avoiding
a risk of inclusion of virus, etc., albumin as a product of such gene
recombination is more preferred.
As used herein, the term "corneal and conjunctival lesion" includes corneal
and conjunctival lesion caused by pathogenic factors such as
keratoconjunctivitis sicca (dry eye), various keratoconjunctivitis,
allergy and infection of microorganisms (e.g. virus, bacteria, fungus,
etc.), chemical factors such as cytotoxicity by chemicals and corrosion
due to acid and alkaline, physical factors such as xerophthalmia, injury
due to foreign matter (e.g. contact lens, etc.) and hot water, and the
like, antiseptics contained in an ophthalmic composition (e.g.
benzalkonium chloride, chlorobutanol, etc.) and ophthalmic agents (e.g.
aminoglycoside antibiotics, non-steroidal anti-inflammatory drugs, IDU,
pimaricin, etc.); defects of ectocornea; corneal erosion; corneal ulcer,
and the like.
As used herein, the term "dry eye" includes not only simple dry eye (tear
decrement) defined as "a condition with decrease or change in quality of
tear, irrespective of the presence or absence of corneal and conjunctival
lesion" but also various dry eye condition such as alacrima,
xerophthalmia, Sjogren syndrome, dry keratoconjunctivitis, Stevens Johnson
syndrome and ocular pemphigoid, blepharitis. Further, the term "dry eye"
includes dry eye after cataract operation and that accompanied with
allergic conjunctivitis, as well as dry eye like condition such as a tear
decrement of VDT (Visual Display Terminal) worker and a tear decrement
without any systemic symptom caused by, for example, dry room due to air
conditioning.
As used herein, the term "treatment" or "treating" refers to any means of
control of the conditions, including prevention, cure and relief of the
conditions and arrestation or relief of development of the condition.
The inventor further found that albumin increases eye surface epithelium
mucin secretion. Therefore, the present invention further provides a
pharmaceutical composition for increasing eye surface mucin secretion
comprising albumin as an active ingredient.
The inventor further found that albumin has a surfactant activity to
diffuses oils. It is suggested that such surfactant activity of albumin
contribute to eye surface tear film stabilization.
The pharmaceutical composition of the present invention may be in a dosage
forms such as tablets, pills, powders, suspensions, capsules,
suppositories, injection preparations, ointments, eye drops, and the like.
It is particularly preferred to locally administer eye drops.
In case of the composition of the present invention is formulated as eye
drops, the composition may contain albumin in an amount of about 1 to 1000
mg/ml, more preferably about 10 to 1000 mg/ml, further preferably about 50
to 1000 mg/ml. The composition may further contain a pharmaceutically
acceptable diluent.
As used herein, the "pharmaceutically acceptable diluent" may be any
diluent which is used for ophthalmic composition known to persons skilled
in the art, for example, water, physiological saline, artificial tear
solution, and the like.
The pharmaceutical composition of the present invention may further
comprise various components that used in a normal ophthalmic composition,
such as stabilizers, sterilizers, buffering agents, isotonic agents,
chelating agents, pH adjusters, surfactants, and the like.
Examples of the stabilizer include normal L-type amino acids such as
glycine and alanine, and the like, monosaccharides such as glucose and
mannose, and the like, disaccharides such as sucrose and maltose, and the
like, sugar alcohols such as mannitol and xylitol, and the like, and
polysaccharides such as dextran, and the like.
Examples of the sterilizer include benzalkonium salt, chlorhexidine salt
and ester of paraoxybenzoate, and the like.
Examples of the buffering agent include boric acid, phosphoric acid, acetic
acid, and citric acid or a salt thereof.
Examples of the isotonic agent include sodium chloride, potassium chloride
and saccharides, and the like.
Examples of the chelating agent include sodium edetate and citric acid, and
the like.
Since it is an ophthalmic composition, pH is preferably adjusted from 5 to
8.
The composition may be administered in a dosage of about 1 to 100
.mu.l/eye, preferably about 10 to 50 .mu.l/eye, and more preferably about
30-50 .mu.l/eye.
In an another aspect, the present invention also provides a use of albumin
for manufacture of a pharmaceutical composition of the present invention.
In still further aspect, the present invention provides a method for
treatment of corneal and conjunctival lesion, which comprises
administering, to a subject in need of such treatment, albumin in an
amount effective.
As used herein, the term "a subject in need of such treatment of corneal
and conjunctival lesion" includes both of a patient who is actually
suffered from corneal and conjunctival lesion and a patient suspected to
be suffered from such lesion. It includes not only the patient whose
corneal and conjunctival lesion has been actually recognized but also the
patient who is suspected of corneal and conjunctival lesion and the
patient in the state where a high possibility of occurring the condition
such as a patient after keratoplasty.
In still further aspect, the present invention provides a method for
treatment of dry eye, which comprises administering, to a subject in need
of such treatment, albumin in an amount effective. As used herein, the
term "a subject in need of such treatment of dry eye" includes both of a
patient who has the dry eye condition and a patient who suspected to be
suffered from dry eye.
In these methods of the present invention, albumin may be the same as that
employed in the above-described composition.
In these methods of the present invention, albumin may be administrated as
the pharmaceutical composition of the present invention. The
administration route is not limited but topical administration to eye is
most preferable.
In these methods of the present invention, "the effective amount" of
albumin, which is an amount for the desirable treatment, may be selected
an optimum according to the patient's symptoms, age, sex, body weight,
diet, other drugs used in combination and various factors which are
recognized by persons skilled in the medical field. This effective amount
may also vary depending on kind or activity of albumin, in addition to the
above factors. Determination of the effective amount is an operation,
which is usually conducted by persons skilled in the art of the medical
field.
In these methods of the present invention, the pharmaceutical composition
may be administered in a dosage of about 1 to 100 .mu.l/eye, preferably
about 10 to 50 .mu./eye and more preferably about 30 to 50 .mu./eye, about
1 to 20 times per day and more preferably, about 5 to 10 times per day, it
is not intended to limit the scope of the invention.
In these methods of the present invention, the artificial tear solution,
which has hitherto been used for treating corneal and conjunctival lesion
or dry eye, may be administered together with albumin. In such a case, the
artificial tear solution may be administered in the amount and schedule as
usual.
In still further aspect of the present invention, the present invention
provides a method for increase of eye surface epitherium mucin secretion,
which comprises administering, to a subject in need of such
administration, albumin in an amount effective. According to the present
method, mucin secretion of the eye surface epithelium may be increased,
and thereby, eye surface tear film may be stabilized.
As used herein, the "a subject in need of such administration" includes
both of a patient with abnormal eye surface mucin secretion and a patient
suspected to have abnormal eye surface mucin secretion due to deficient of
eye surface epithelium, such as a patient of dry eye or corneal and
conjunctival lesion.
In this method of the present invention, albumin may be administrated as
the pharmaceutical composition of the present invention. The
administration route is not limited but topical administration to eye is
most preferable.
In this method of the present invention, "the effective amount" of albumin,
which is an amount for increase eye surface mucin secretion, may be
selected an optimum amount according to the patient's symptoms, age, sex,
body weight, diet, other drugs used in combination and various factors
which are recognized by persons skilled in the medical field. This
effective amount may also vary depending on kind or activity of albumin,
in addition to the above factors. Determination of the effective amount is
an operation, which is usually conducted by persons skilled in the art of
the medical field.
In this method of the present invention, the pharmaceutical composition may
be administered in a dosage of about 1 to 100 .mu.l/eye, preferably about
10 to 50 .mu.l/eye and more preferably about 30 to 50 .mu.l/eye, about 1
to 20 times per day and more preferably, about 5 to 10 times per day, it
is not intended to limit the scope of the invention.
In this method of the present invention, the artificial tear solution,
which has hitherto been used for treating corneal and conjunctival lesion
or dry eye, may be administered together with albumin. In such a case, the
artificial tear solution may be administered in the amount and schedule as
usual.
FORMULATION EXAMPLE
In the examples of this application, donated blood albumin preparations
manufactured by The Green Cross Corporation (Osaka, Japan) were used.
These albumin preparations were obtained by using an albumin fraction
which was isolated/purified plasma of a blood donar as a raw material
according to a Cohn's cold ethanol fractionation method, preparing
according to the following Formulation Example 1 and 2, and heat-treating
at 60.degree. C. for 10 hours.
Formulation Example 1
______________________________________
(Albumin content: 5%)
______________________________________
Human serum albumin
50 mg/ml
Acetyltriptophan sodium 1.07 mg/ml
Sodium caprylate 0.66 mg/ml
______________________________________
Formulation Example 2
______________________________________
(Albumin content: 25%)
______________________________________
Human serum albumin
250 mg/ml
Acetyltriptophan sodium 5.37 mg/ml
Sodium caprylate 3.32 mg/ml
______________________________________
Test Example 1
Three dry eye patients with corneal and conjunctival lesion (female aged
64, female aged 61 and female aged 34) were administrated to their eyes
with 25% human serum albumin (albumin content: 250 mg/ml) of Formulation
Example 1, 10 times per day with a dosage of 30 to 50 .mu.l/eye. Together
with administration of albumin, an artificial tear solution was also
administered to the eyes.
An intravital stain examination was conducted before and after the
administation to estimate the degree of corneal and conjunctival lesion.
As the intravital stain examination, rose bengal staining and fluorescein
staining were conducted. The rose bengal staining (RB) provides an index
of corneal and conjunctival lesion, and scoring was conducted by
estimating the degree of staining of nasal and aural sides of bulbar
conjunctiva and cornea by a score of 0-3 (total scores of 9). The
fluorescein staining (F) provides an index of corneal lesion, and scoring
was conducted by estimating the degree of corneal lesion by a score of
0-3. In both cases, scoring was conducted according to a van Bijsterreld's
evaluation method. In both cases, the higher the numerical value is, the
more severe the lesion is. The results are shown in Table 1.
TABLE 1
__________________________________________________________________________
Effect of albumin administration
Female aged
Female aged
sixty-four sixty-one
Patient 14 Days 14 days Female aged
thirty-four*
Administration period
Right eye
Left eye
Right eye
Left eye
7 days
__________________________________________________________________________
RB
Before administration 7 7 8 8 The
superior limbic part is
strongly stained.
After administration 3 3 5 5 The
superior limbic part is
scarcely stained.
F
Before administration 2 2 3 2 The
superior limbic part is
strongly stained.
After administration 1 1 1 1 The
superior limbic part is
scarcely stained.
__________________________________________________________________________
*She was suffered from superior limbic keratoconjunctivitis in addition t
the dry eye.
Test Example 2
Male aged 62 with defects of ectocornea after keratoplasty
After penetrating keratoplasty of the right eye, defects of ectocornea were
continued and 0.1% Hyaleinmini (trademark) (containing 0.1% of hyaluronic
acid) was administered. However, no effect was observed and, therefore,
administration of 0.1% Hyaleinmini was terminated. Thereafter, 25% human
serum albumin (albumin content: 250 mg/ml) of Formulation Example 1 was
administered to the eyes 10 times per day with a dosage of 30 to 50
.mu.l/eye for one week. One week after the beginning of administration,
ectocornea is formed and defects of ectocornea were improved.
Test Example 3
Female aged 64 with defects of ectocornea after keratoplasty
After penetrating keratoplasty of the left eye, limbus transplantation and
amnion transplantation, defects of ectocornea were detected by the
fluorescein staining. To the eyes of this patient, 5% human serum albumin
(albumin content: 50 mg/ml) of Formulation Example 2 was administered 10
times per day with a dosage of 30 to 50 .mu.l/eye for four weeks. Two
weeks after the beginning of the administration, fluorescein staining
showed improvement in defects of ectocornea. Four weeks after the
beginning of the administration, no staining was observed and an apparent
improvement in defects of ectocornea was recognized.
Test Example 4
Female aged 72 with Sjogren syndrome
To the eyes of the patient with defects of ectocornea accompanied with
Sjogren syndrome, Intal (trademark) (containing sodium cromoglicate), 0.1%
Flumetholon (trademark) (containing fluorometholone) and an artificial
tear solution were administered. However, no improvement in defects of
ectocornea was observed and the administration was terminated. Then, to
the eyes of the patient, 5% human serum albumin (albumin content: 50
mg/ml) of Formulation Example 2 was administered 6 times per day with a
dosage of 30 to 50 .mu.l/eye for four weeks. Four weeks after the
beginning of the administration, defects of ectocornea were not observed.
Test Example 5
Female aged 45 with Sjogren syndrome
To the eyes of the patient with corneal and conjunctival lesion accompanied
with Sjogren syndrome, 5% human serum albumin (albumin content: 50 mg/ml)
of Formulation Example 2 was administered 10 times per day with a dosage
of 30 to 50 .mu.l/eye for four weeks.
Before and after the administration, an intravital stain examination (rose
bengal staining and fluorescein staining) was conducted to estimate the
degree of corneal and conjunctival lesion. The intravital stain
examination was conducted by applying 2 .mu.l of a mix solution containing
1% rose bengal and 1% fluorescein to the lower eyelids of the patient
using a micro-pipette accurately, making the patient to blink several
times, and then observing the eyes. Rose bengal staining (RB) was measured
with white light of slit lamp and fluorescein staining (F) was measured
with cobalt blue light. The extent of staining was scored from 0 to 9. The
results are shown in Table 2.
TABLE 2
______________________________________
Effects of albumin administration
After administration
before administration (four weeks)
______________________________________
RB score
right eye 7 2
left eye 7 2
F score right eye 9 2
left eye 9 2
______________________________________
Test Example 6
Three dry eye patients (female aged 64, female aged 61 and female aged 34)
were administrated with 25% human serum albumin (albumin content: 250
mg/ml) of Formulation Example 1 to their eyes 10 times per day with a
dosage of 30 to 50 .mu.l/eye. Together with administration of albumin, an
artificial tear solution was also administrated to the eye.
Before and after the administration, subjective symptoms of the patients
were estimated and scored. The subjective symptom includes eye ache, eye
dry feeling and eye foreign body feeling. Estimation of the symptoms was
made by the patients themselves. The worst subjective symptom was scored
as 100 point and best or normal as 0. The results are shown in Table 3.
TABLE 3
______________________________________
Improvement of subjective symptoms by administration of albumin
Patient Female, aged 64
Female, aged 61
Female,
term of 14 days 14 days aged 34*
administration
right eye
left eye
right eye
left eye
7 days
______________________________________
before 100 100 100 100 100
administration
after 0 30 50 50 20
administration
______________________________________
*: she was suffering from superior limbic keratoconjunctivitis in additio
to the dry eye.
Test Example 7
To the eyes of a dry eye patient accompanying with Sjogren syndrome (female
aged 74), 5% human serum albumin (albumin content: 50 mg/ml) of
Formulation Example 2 was administered 6 times per day with a dosage of 30
to 50 .mu.l/eye for eight weeks.
After eight weeks, subjective symptoms including eye ache, eye dry feeling
and eye foreign body feeling were almost perfectly removed. Administration
of albumin was stopped.
Test Example 8
The ability of albumin to increase mucin secretion of eye epitherium was
investigated.
CCL cells (conjunctival epitherium cell strain) were cultured in TCM199
medium (GIBCO) containing 10% (w/v) human serum albumin (SIGMA) for 24
hours according to conventional cell culture condition. As a culture
control, the cells were cultured with the TCM 199 medium without human
serum albumin.
The cultured cells were harvested from the culture vessel using
trypsin-EDTA, fixed with paraformaldehyde for 30 minutes and washed with
phosphate buffered saline (PBS) three times. The obtained cells were
blocked (4.degree. C., 30 min.) with normal goat serum and the blocked
cells were reacted (4.degree. C., 30 min.) with mouse anti-mucin antibody
(Muc 1), and then washed with PBS three times. The cells further reacted
(4.degree. C., 30 min.) with FITC-labeled anti-mouse IgG antibody and then
washed three times with PBS.
The obtained cells were measured with Epics (Colter Co.) by flow-cytometry
method to determine the proportion (%) of mucin generating cells (positive
cells) to the whole cells. The result is shown in Table 4.
TABLE 4
______________________________________
increase of mucin secretion by albumin
mucin generating cells (positive)
______________________________________
control (without albumin)
15.2%
cultured with 10% albumin 35.5%
______________________________________
According to the result, addition of albumin to the culture medium
apparently increased mucin secretion ability of conjunctival epitherium.
Test Example 9
Ability to diffuse oil by albumin:
10 vol % of caster oil was added to an artificial tear solution. The oil
and the solution separated out and oil drops were formed. In this system,
10% (v/v of artificial tear solution) of 5% albumin solution was added,
the oil diffused over the surface of the water phase and the oil drops
disappeared.
According to this example, it is suggested that albumin can act as a
surfactant to protect vaporization of tear solution from eye surface.
INDUSTRIAL APPLICABILITY
The pharmaceutical composition, method and use of the present invention are
useful for treatment of corneal and conjunctival lesion and dry eye. The
pharmaceutical composition, method and use of the present invention are
also useful for increasing mucin secretion of eye surface epitherium.
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